The efficacy of a vaccine can be enhanced by the addition of adjuvants, which are substances that increase the magnitude, and/or influence the nature of the immune response to a foreign antigen. Due to safety concerns, very few adjuvants have been approved for use in human vaccines. Consequently, efforts have focused on exploiting the properties of "natural adjuvants," which are innate immunostimulatory components of the vaccinated individual. One natural adjuvant that has significant potential for improving vaccine efficacy is the dendritic cell (DC). DCs play a central role in initiating innate and adaptive immune responses to foreign antigens;and therefore, targeting vaccines to DCs is a rational strategy for enhancing vaccine efficacy. Our long-term goal is to develop a broadly applicable strategy for targeting vaccines to DCs in vivo. The objective of this proposal is to develop and evaluate a DC-targeting vaccine strategy based on a bacterial protein called protein L (PpL). PpL is an immunoglobulin (Ig)-binding protein that binds to the variable domain of Ig kappa light chains derived from human, non-human primates and the mouse. We have shown that vaccine vectors and vector-expressed antigens containing appropriate PpL sequences spontaneously form immune complexes (ICs) with non-immune Ig in vitro. If this phenotype is reproduced in vivo, then the vector- or antigen-containing ICs should be targeted to DCs via binding to Fc receptors (FcR), and these interactions should result in enhanced immune responses to the PpL-modified vector/antigens. In this project we will study the immunological consequences of attaching PpL sequences to a replicating viral vaccine vector (Sindbis virus) and to model vector-expressed vaccine antigens. The central hypothesis of this project is the attachment of PpL sequences to viral vaccine vectors and vector-expressed antigens will enhance their immunogenic properties by facilitating their interactions with DCs in vivo. This hypothesis will be tested by pursuing the following two specific aims: (1) Determine if incorporation of one or more PpL Ig-binding domain(s) into Sindbis virus vectors and vector-expressed antigens enhances their interactions with primary DCs grown in culture and in vivo. (2) Characterize the cellular and humoral immune responses in mice to native and PpL-modified viral vector proteins and vector-expressed vaccine antigens following intranasal, intradermal, and subcutaneous immunizations.